Ethanol (EtOH) inhibition of large conductance, Ca⁺⁺-activated K⁺ (BK_Ca) channel activity in aortic myocytes may contribute to the direct contraction of aortic smooth muscle produced by acute exposure to alcohol. In this tissue, BK_Ca channels consist of pore forming (, encoded by the slo gene) and modulatory (₁) subunits. Here, modulation of aortic smooth muscle BK_Ca channels by acute alcohol was explored by expressing bslo subunits in Xenopus oocytes, in the absence and presence of ₁ subunits, and studying channel responses to clinically relevant concentrations of EtOH in excised membrane patches. Overall, average values of bslo channel activity NP_o)in response to acute EtOH (3-200 mM) mildly decrease when compared to pre-EtOH, isosmotic control values. However, data show qualitative heterogeneity in bslo channel responses to EtOH at all concentrations tested. In the majority of patches (42 out of 71 patches, i.e., 59%) a reversible reduction in NP_o is observed. In this subset, the maximal effect is obtained with 100 mM EtOH, at which NP_o reaches 46.2±9% of control. The presence of ₁ subunits, which determines channel sensitivity to dihydrosoyaponin-I and 17-estradiol, fails to modify EtOH action on bslo channels. EtOH reduction of bslo channel activity results from a marked increase in the mean closed time. While the voltage-dependence of gating remains largely unaffected, the apparent effectiveness of Ca⁺⁺ to gate the channel is decreased by EtOH. These changes occur in the absence of modifications of channel conduction properties. In conclusion, a new molecular mechanism that may contribute to EtOH-induced aortic smooth muscle contraction has been identified and characterized: a functional interaction between EtOH and the bslo subunit and/or its lipid microenvironment, which leads to a decrease in BK_Ca channel activity.